Ligand-receptor interactions are essential to a number of cell signaling pathways. Growth factors, cytokines and other regulatory proteins use these interactions to mediate cell responses. Proteins that inhibit or facilitate these processes have potential as therapeutics.
Given some of the drawbacks of monoclonal antibody approaches to inhibiting ligand-receptor functions such as expensive manufacturing, large size, limited penetration into tissues and undesirable side effects, researchers have been focusing on the use of non-antibody proteins as therapeutic agents. Furthermore, therapeutic antibody strategies are generally limited to inhibiting, or antagonizing, a signaling pathway and not competent to strategies to enhance, or agonize, a pathway. Thus, new protein engineering approaches are being explored to develop ligands and receptors as agonists and antagonists of clinically important targets as an alternative to antibody strategies.
Circular permutation involves the linking of the native amino and carboxy ends of a protein, generally with a linker, and creating new amino and carboxy termini by cleaving at a new site within the protein sequence, generally a loop; such that the primary sequence of the resulting protein is reordered, while the secondary structure (and activity) is retained. Thus, creation of the new termini may provide better locations for attachment of a fusion partner relative to the native termini.
Circular permutation of a protein ligand provides a means by which a protein may be altered to produce new carboxyl and amino termini without diminishing the specificity and binding affinity of the altered protein ligand for its target relative to its native form. Additionally, the new termini can be preferentially moved to a location preferential for incorporating the circularly permuted ligand into a fusion polypeptide, and demonstrate better activity compared with a fusion polypeptide containing the native (non-circularly permuted) ligand.
The present invention provides fusion polypeptides comprising ligands modified by circular permutation which function as agonists, super agonists or antagonists of a signaling pathway. Such fusion polypeptides are beneficial in the treatment of many disorders, conditions, and diseases that rely on ligand-receptor interaction and signal transduction. For example, such fusion polypeptides that act as antagonists of a target receptor have potential as therapeutics for cancer and autoimmune disorders. Such fusion polypeptides that act as agonists or superagonists of a signaling pathway have the potential, for example, in cancer or regenerative medicine.